In little more than 16 years, HIV and its associated illness, AIDS, has become one of the largest plagues to affect humankind. Throughout the world it is estimated that there are 40 million people infected with HIV. Health officials predict an additional 60 million people will become infected by 2010. In addition to reverse transcriptase and protease, integrase has become a highly sought target for directing antivirals to treat individuals with AIDS. The proposed study will offer valuable insight into the mechanism of retroviral integration, and that a more complete understanding of the mechanism will provide useful information for those concerned with designing effective inhibitors of the retroviral integrase. In effect, the substrate analogs used herein may offer insight into the chemistry and architecture of the interaction of the viral DNA substrate and integrate, and how it can be blocked. Our major effort will be to build a detailed biochemical picture of the catalytic site of HIV-1 integrase and its interactions with viral end DNA substrates. The reasons catalyzed by the retroviral integrase can be divided into two major steps: 3'-processing and strand transfer. One of the hypothesis proposed in this research suggests these steps can be broken down further into additional discrete operational units such as binding, fraying, cleavage, target acquisition, and transesterification. In the context of these operational units, it is proposed that the interaction of integrase with the viral end DNA (and then later with t he target DNA) requires substantial changes in the conformation of the protein structure. To test this hypothesis, experiments were designed using novel substrates which will block the ability of integrase to mediate the following steps: fraying, 3-processing, and strand transfer. This approach allows for the assembly of discrete complexes along the integration pathway. The "dead-end" viral end substrates will allow studies to probe the architecture of three discrete integrase-DNA complexes, and map the conformation changes, amino acid contacts and multimeric states that occur at specific steps in the integration reaction mechanism. These complexes will be characterized using standard biochemical techniques, which will provide "free-frame" images of integrase with its substrates.